Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Article
  • Published:

A vesicle-trafficking protein commandeers Kv channel voltage sensors for voltage-dependent secretion

Nature Plants volume 1, Article number: 15108 (2015) Cite this article

Subjects

An Erratum to this article was published on 07 October 2015

Abstract

Growth in plants depends on ion transport for osmotic solute uptake and secretory membrane trafficking to deliver material for wall remodelling and cell expansion. The coordination of these processes lies at the heart of the question, unresolved for more than a century, of how plants regulate cell volume and turgor. Here we report that the SNARE protein SYP121 (SYR1/PEN1), which mediates vesicle fusion at the Arabidopsis plasma membrane, binds the voltage sensor domains (VSDs) of K+ channels to confer a voltage dependence on secretory traffic in parallel with K+ uptake. VSD binding enhances secretion in vivo subject to voltage, and mutations affecting VSD conformation alter binding and secretion in parallel with channel gating, net K+ concentration, osmotic content and growth. These results demonstrate a new and unexpected mechanism for secretory control, in which a subset of plant SNAREs commandeer K+ channel VSDs to coordinate membrane trafficking with K+ uptake for growth.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: KC1 K+ channel interaction with SYP121 requires the conserved RYxxWE motif of the voltage sensor domain.
Figure 2: Secretory traffic rescue by the KC1 VSD depends on the RYxxWE motif and the native SYP121.
Figure 3: VSD mutants VSDF129W and VSDD132E alter the voltage-sensitivity of VSD conformation, SYP121 binding, and secretory traffic rescue.
Figure 4: KC1 and its VSD promote secretion in parallel with VSD voltage bias.
Figure 5: KC1, its VSD, and SYP121ΔC suppress plant growth and, differentially, K+ and osmotic content.
Figure 6: Governor model for SYP121 and Kv channel interaction and their uncoupling by SYP121ΔC, VSDwt and the VSDD132E mutant.

Similar content being viewed by others

References

  1. Zonia, L. & Munnik, T. Life under pressure: hydrostatic pressure in cell growth and function. Trends Plant Sci. 12, 90–97 (2007).

    Article  CAS  PubMed  Google Scholar 

  2. Walter, A., Silk, W. K. & Schurr, U. Environmental effects on spatial and temporal patterns in leaf and root growth. Annu. Rev. Plant Biol. 60, 279–304 (2009).

    Article  CAS  PubMed  Google Scholar 

  3. Orlowski, J. & Grinstein, S. Emerging roles of alkali cation/proton exchangers in organellar homeostasis. Curr. Opin. Cell Biol. 19, 483–492 (2007).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Amtmann, A. & Blatt, M. R. Regulation of macronutrient transport. New Phytol. 181, 35–52 (2009).

    Article  CAS  PubMed  Google Scholar 

  5. Grefen, C., Honsbein, A. & Blatt, M. R. Ion transport, membrane traffic and cellular volume control. Curr. Opin. Cell Biol. 14, 332–339 (2011).

    Article  CAS  Google Scholar 

  6. Scheible, W. R. et al. An Arabidopsis mutant resistant to thaxtomin A, a cellulose synthesis inhibitor from Streptomyces species. Plant Cell 15, 1781–1794 (2003).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Geelen, D. et al. The abscisic acid-related SNARE homolog NtSyr1 contributes to secretion and growth: evidence from competition with its cytosolic domain. Plant Cell 14, 387–406 (2002).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Eisenach, C., Chen, Z. H., Grefen, C. & Blatt, M. R. The trafficking protein SYP121 of Arabidopsis connects programmed stomatal closure and K+ channel activity with vegetative growth. Plant J. 69, 241–251 (2012).

    Article  CAS  PubMed  Google Scholar 

  9. Blatt, M. R. Cellular signaling and volume control in stomatal movements in plants. Annu. Rev. Cell Dev. Biol. 16, 221–241 (2000).

    Article  CAS  PubMed  Google Scholar 

  10. Galvan-Ampudia, C. S. & Testerink, C. Salt stress signals shape the plant root. Curr. Opin. Cell Biol. 14, 296–302 (2011).

    Article  CAS  Google Scholar 

  11. Campanoni, P. & Blatt, M. R. Membrane trafficking and polar growth in root hairs and pollen tubes. J. Exp. Bot. 58, 65–74 (2007).

    Article  CAS  PubMed  Google Scholar 

  12. Lipka, V., Kwon, C. & Panstruga, R. SNARE-Ware: the role of SNARE-Domain proteins in plant biology. Annu. Rev. Cell Dev. Biol. 23, 147–174 (2007).

    Article  CAS  PubMed  Google Scholar 

  13. Jahn, R. & Scheller, R. H. SNAREs - engines for membrane fusion. Nature Rev. Mol. Cell Biol. 7, 631–643 (2006).

    Article  CAS  Google Scholar 

  14. Grefen, C. & Blatt, M. R. SNAREs - molecular governors in signalling and development. Curr. Opin. Cell Biol. 11, 600–609 (2008).

    Article  CAS  Google Scholar 

  15. Leyman, B., Geelen, D., Quintero, F. J. & Blatt, M. R. A tobacco syntaxin with a role in hormonal control of guard cell ion channels. Science 283, 537–540 (1999).

    Article  CAS  PubMed  Google Scholar 

  16. Collins, N. C. et al. SNARE-protein-mediated disease resistance at the plant cell wall. Nature 425, 973–977 (2003).

    Article  CAS  PubMed  Google Scholar 

  17. Blatt, M. R., Grabov, A., Brearley, J., HammondKosack, K. & Jones, J. D. G. K+ channels of Cf-9 transgenic tobacco guard cells as targets for Cladosporium fulvum Avr9 elicitor-dependent signal transduction. Plant J. 19, 453–462 (1999).

    Article  CAS  PubMed  Google Scholar 

  18. Dangl, J. L. & Jones, J. D. G. Plant pathogens and integrated defence responses to infection. Nature 411, 826–833 (2001).

    Article  CAS  PubMed  Google Scholar 

  19. Honsbein, A., Blatt, M. R. & Grefen, C. A molecular framework for coupling cellular volume and osmotic solute transport control. J. Exp. Bot. 62, 2363–2370 (2011).

    Article  CAS  PubMed  Google Scholar 

  20. Honsbein, A. et al. A tripartite SNARE-K+ channel complex mediates in channel-dependent K+ nutrition in Arabidopsis. Plant Cell 21, 2859–2877 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Grefen, C. et al. A novel motif essential for SNARE interaction with the K+ channel KC1 and channel gating in Arabidopsis. Plant Cell 22, 3076–3092 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Sanderfoot, A. Increases in the number of SNARE genes parallels the rise of multicellularity among the green plants. Plant Physiol. 144, 6–17 (2007).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Dreyer, I. & Blatt, M. R. What makes a gate? The ins and outs of Kv-like K+ channels in plants. Trends Plant Sci. 14, 383–390 (2009).

    Article  CAS  PubMed  Google Scholar 

  24. Grefen, C. & Blatt, M. R. A 2in1 cloning system enables ratiometric bimolecular fluorescence complementation (rBiFC). Biotechniques 53, 311–314 (2012).

    Article  CAS  PubMed  Google Scholar 

  25. Chakrapani, S., Cuello, L. G., Cortes, D. M. & Perozo, E. Structural dynamics of an isolated voltage-sensor domain in a lipid bilayer. Structure 16, 398–409 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Palovcak, E., Delemotte, L., Klein, M. L. & Carnevale, V. Evolutionary imprint of activation: the design principles of VSDs. J. Gen. Physiol. 143, 145–156 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Richter, S., Voss, U. & Juergens, G. Post-Golgi traffic in plants. Traffic 10, 819–828, (2009).

    Article  CAS  PubMed  Google Scholar 

  28. Tyrrell, M. et al. Selective targeting of plasma membrane and tonoplast traffic by inhibitory (dominant-negative) SNARE fragments. Plant J. 51, 1099–1115 (2007).

    Article  CAS  PubMed  Google Scholar 

  29. Karnik, R. et al. Arabidopsis Sec1/Munc18 protein sec11 is a competitive and dynamic modulator of SNARE binding and SYP121-dependent vesicle traffic. Plant Cell 25, 1368–1382 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Hecker, A. et al. Binary 2in1 vectors improve in planta (co-) localisation and dynamic protein interaction studies. Plant Physiol. 168,776–787 (2015).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  31. Sutter, J. U., Campanoni, P., Tyrrell, M. & Blatt, M. R. Selective mobility and sensitivity to SNAREs is exhibited by the Arabidopsis KAT1 K+ channel at the plasma membrane. Plant Cell 18, 935–954 (2006).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. Karnik, R. et al. Binding of SEC11 indicates its role in SNARE recycling after vesicle fusion and identifies two pathways for vesicular traffic to the plasma membrane. Plant Cell 27, 675–694 (2015).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  33. Assaad, F. F. et al. The PEN1 syntaxin defines a novel cellular compartment upon fungal attack and is required for the timely assembly of papillae. Mol. Biol. Cell 15, 5118–5129 (2004).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. Zhang, Z. G. et al. A SNARE-protein has opposing functions in penetration resistance and defence signalling pathways. Plant J. 49, 302–312 (2007).

    Article  CAS  PubMed  Google Scholar 

  35. Rehman, R. U. et al. Tomato Rab11a characterization evidenced a difference between SYP121-dependent and SYP122-dependent exocytosis. Plant Cell Physiol. 49, 751–766 (2008).

    Article  CAS  PubMed  Google Scholar 

  36. Fujiwara, M. et al. Interactomics of Qa-SNARE in Arabidopsis thaliana. Plant Cell Physiol. 55, 781–789 (2014).

    Article  CAS  PubMed  Google Scholar 

  37. Latorre, R. et al. Molecular coupling between voltage sensor and pore opening in the Arabidopsis inward rectifier K+ channel KAT1. J. Gen. Physiol. 122, 459–469 (2003).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Lai, H. C., Grabe, M., Jan, Y. N. & Jan, L. Y. The S4 voltage sensor packs against the pore domain in the KAT1 voltage-gated potassium channel. Neuron 47, 395–406 (2005).

    Article  CAS  PubMed  Google Scholar 

  39. Lefoulon, C. et al. Voltage-sensor transitions of the inward-rectifying K+ channel KAT1 indicate a closed-to-open latching mechanism that is biased by hydration around the S4 alpha-helix. Plant Physiol. 166, 960–975, (2014).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  40. Tsutsui, H., Karasawa, S., Okamura, Y. & Miyawaki, A. Improving membrane voltage measurements using FRET with new fluorescent proteins. Nature Methods 5, 683–685 (2008).

    Article  CAS  PubMed  Google Scholar 

  41. Youvan, D. C. et al. Calibration of fluorescence resonance energy transfer in microscopy using genetically engineered GFP derivatives on nickel chelating beads. Biotechnology 3, 1–18 (1997).

    Google Scholar 

  42. Chen, Z. H., Grefen, C., Donald, N., Hills, A. & Blatt, M. R. A bicistronic, Ubiquitin-10 promoter-based vector cassette for transient transformation and functional analysis of membrane transport demonstrates the utility of quantitative voltage clamp studies on intact Arabidopsis root epidermis. Plant Cell Environ. 34, 554–564 (2011).

    Article  CAS  PubMed  Google Scholar 

  43. Fricker, M. D., Runions, J. & Moore, I. Quantitative fluorescence microscopy: from art to science. Ann. Rev. Plant Biol. 57, 79–107 (2006).

    Article  CAS  Google Scholar 

  44. Sokolovski, S., Hills, A., Gay, R. & Blatt, M. R. Functional interaction of the SNARE protein NtSyp121 in Ca2+ channel gating, Ca2+ transients and ABA signalling of stomatal guard cells. Mol. Plant 1, 347–358 (2008).

    Article  CAS  PubMed  Google Scholar 

  45. Wang, Y., Hills, A. & Blatt, M. R. Systems analysis of guard cell membrane transport for enhanced stomatal dynamics and water use efficiency. Plant Physiology 164, 1593–1599 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  46. Wang, Y. et al. Overexpression of plasma membrane H+-ATPase in guard cells promotes light-induced stomatal opening and enhances plant growth. Proc. Natl Acad. Sci. USA 111, 533–538 (2014).

    Article  CAS  PubMed  Google Scholar 

  47. Zhang, B. et al. The R-SNARE VAMP721 interacts with KAT1 and KC1 K+ channels to moderate K+ current at the plasma membrane. Plant Cell 27, 1697–1717 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  48. Leung, Y. M., Kwan, E. P., Ng, B., Kang, Y. & Gaisano, H. Y. SNAREing voltage-gated K+ and ATP-Sensitive K+ channels Tuning beta-Cell excitability with syntaxin-1A and other exocytotic proteins. Endocrine Rev. 28, 653–663 (2007).

    Article  CAS  Google Scholar 

  49. Iwasaki, H. et al. A voltage-sensing phosphatase, Ci-VSP, which shares sequence identity with PTEN, dephosphorylates phosphatidylinositol 4,5-bisphosphate. Proc. Natl Acad. Sci. USA 105, 7970–7975 (2008).

    Article  CAS  PubMed  Google Scholar 

  50. Tombola, F., Ulbrich, M. H. & Isacoff, E. Y. The voltage-gated proton channel Hv1 has two pores, each controlled by one voltage sensor. Neuron 58, 546–556 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  51. Lawson, T. & Blatt, M. R. Stomatal size, speed, and responsiveness impact on photosynthesis and water use efficiency. Plant Physiol. 164, 1556–1570 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  52. Grefen, C. et al. A ubiquitin-10 promoter-based vector set for fluorescent protein tagging facilitates temporal stability and native protein distribution in transient and stable expression studies. Plant J. 64, 355–365 (2010).

    Article  CAS  PubMed  Google Scholar 

  53. Karnik, A., Karnik, R. & Grefen, C. SDM-Assist software to design site-directed mutagenesis primers introducing “silent” restriction sites. BMC Bioinformatics 14, 105 (2013).

    Article  PubMed  PubMed Central  Google Scholar 

  54. Clough, S. J. & Bent, A. F. Floral dip: a simplified method for Agrobacterium-mediated transformation of Arabidopsis thaliana. Plant J. 16, 735–743 (1998).

    Article  CAS  Google Scholar 

  55. Grefen, C. & Blatt, M. R. Do calcineurin B-like proteins interact independently of the serine threonine kinase CIPK23 with the K+ channel AKT1? Lessons learned from a menage a trois. Plant Physiol. 159, 915–919 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  56. Blatt, M. R. & Grefen, C. in Arabidopsis Protocols Vol. 3 (eds Sanchez-Serrano, J. J. & Salinas, J. ) 487–507 (2014).

    Book  Google Scholar 

Download references

Acknowledgements

We are grateful to N. Donald and L. Matas for technical support. G. Boswell and A. Ruiz-Pardo helped with Xenopus and plant maintenance. This work was supported by a Chinese Scholarship Council award to B.Z. and by grants BB/H0024867/1, BB/I024496/1, BB/K015893/1, BBL001276/1 and BB/M001601/1 from the UK Biotechnology and Biological Sciences Research Council to M.R.B. C.G. is supported by an Emmy Noether Fellowship from the Deutsche Forschungsgemeinschaft GR 4251/1-1.

Author information

Author notes

  1. Christopher Grefen

    Present address: † Present address: ZMBP Department of Developmental Genetics, Auf der Morgenstelle 32, Tübingen D72076, Germany,

  2. Rucha Karnik and Michael R. Blatt: These authors jointly supervised this work

  3. Christopher Grefen, Rucha Karnik and Emily Larson: These authors contributed equally to this work.

Authors and Affiliations

  1. Laboratory of Plant Physiology and Biophysics, Bower Building, University of Glasgow, Glasgow, G12 8QQ, UK

    Christopher Grefen, Rucha Karnik, Emily Larson, Cécile Lefoulon, Yizhou Wang, Sakharam Waghmare, Ben Zhang, Adrian Hills & Michael R. Blatt

Authors
  1. Christopher Grefen
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Rucha Karnik
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Emily Larson
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Cécile Lefoulon
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Yizhou Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Sakharam Waghmare
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Ben Zhang
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Adrian Hills
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Michael R. Blatt
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

M.R.B. conceived and wrote the manuscript with C.G., R.K. and E.L.; C.G., designed and generated vectors; C.G., R.K., B.Z. and C.L. prepared constructs; C.G. and B.Z. carried out the split-ubiquitin studies; C.G., R.K. and E.L. generated the stable lines, and R.K. and E.L. analysed the lines; R.K. and S.W. carried out pulldown and immunoblot analysis; M.R.B. carried out the rBiFC, secretion and Mermaid FRET assays; C.L. recorded K+ currents in oocytes, Y.W. recorded K+ currents in Arabidopsis; A.H. developed analysis utilities for this work.

Corresponding author

Correspondence to Michael R. Blatt.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Grefen, C., Karnik, R., Larson, E. et al. A vesicle-trafficking protein commandeers Kv channel voltage sensors for voltage-dependent secretion. Nature Plants 1, 15108 (2015). https://doi.org/10.1038/nplants.2015.108

Download citation

  • Received:

  • Accepted:

  • Published:

  • DOI: https://doi.org/10.1038/nplants.2015.108

This article is cited by

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing